In the production processes for compounds serving as source materials of chemical products, enzymatic reactions are currently used due to their high conversion and selectivity, particularly due to their high stereoselectivity in the case of optically active compounds. For example, enzymatic reactions catalyzed by nitrilase, nitrile hydratase, amidase and the like are used for production of nitrile compounds (e.g., acrylamide, acrylic acid) serving as source materials of various chemical products. Nitrilase is an enzyme converting a nitrile group into a carboxylic acid group through hydrolysis, nitrile hydratase is an enzyme converting a nitrile group into an amido group through hydration, and amidase is an enzyme converting an amido group into a carboxylic acid group through hydrolysis.
Microorganisms having these enzymes have been screened from the natural environment including soil. Techniques commonly used for screening purposes involve enriching microorganisms which grow using nitrile compounds or the like as a sole nitrogen or carbon source, and selecting a microorganism having enzymatic activity from among the resulting microorganisms. For verification of enzymatic activity, a nitrile compound or an amide compound is reacted with each microorganism and the resulting product is analyzed with an instrument for high performance liquid chromatography or gas chromatography, etc.
On the other hand, there is a research report showing that microorganisms which can be isolated and cultured by currently used techniques constitute only less than 1% of the microorganisms present in the natural environment (M. S. Rappe and S. J. Giovannoni, Annu. Rev. Microbiol., 57, 369 (2003)).
For this reason, another procedure (metagenome screening) has been used in recent studies, which involves directly isolating genes (environmental DNAs or metagenomes) from the natural environment, instead of isolating microorganisms as in conventional techniques, and then screening the isolated genes to select a useful enzyme gene. The use of this procedure requires techniques for preparing a very large number of metagenome-derived recombinants and efficiently selecting active recombinants from among these recombinants. However, screening with an instrument for high performance liquid chromatography or gas chromatography is not high-throughput. Thus, there has been a demand for a novel technique which allows high-throughput screening.
Meanwhile, an invention of a fluorescent probe or fluorescent labeling agent is known as an invention directed to a fluorescent substance (WO2004/005917, WO2006/019105). However, the fluorescent substrate of the present invention useful for detection of enzymatic activity is not known.